The present invention is directed to improved methods for preparing L-2-oxothiazolidine-4-carboxylate and its carboxylic acid, namely, L-2-oxothiazolidine-4-carboxylic acid. The present invention improves the separation of phenol from a mixture containing L-2-oxothiazolidine-4-carboxylate. The present method is simpler, less costly, safer to employ, and less toxic than the methods of the prior art.
U.S. Pat. No. 4,647,571 ("the '571 patent"), which issued to Meister et al., teaches that L-2-oxothiazolidine-4-carboxylate is an important compound in the intracellular delivery of L-cysteine in the body. Poisoning can occur by the excessive use of pain-killing drugs, such as those containing the active ingredients N-acetyl-para-aminophenol, paracetamol, and acetaminophen. Such drugs cause the level of glutathione in the liver to decrease. L-2-oxothiazolidine-4-carboxylate increases intracellular levels of L-cysteine (see, Meister, Science 220: 472 (1983)), which, in turn, increases the glutathione level in liver cells, which protects against the toxicity of such poisons.
The '571 patent also discloses that L-cysteine is relatively toxic extracellularly and therefore cannot be administered in significant quantities directly to the subject. See also Karlsen et al., Brain Res. 208: 167 (1981). The '571 patent teaches that by administering L-2-oxothiazolidine-4-carboxylate to the subject, L-cysteine is formed within the cells without the resultant toxicity which occurs when L-cysteine is administered extracellularly.
Either L-2-oxothiazolidine-4-carboxylate (i.e., the neutral salt form) or its carboxylic acid can be directly administered to the patient. For example, the salt is generally used for intravenous administration and the acid for administration by tablet. However, because the pH of blood is about 7, L-2-oxothiazolidine-4-carboxylate will be the active form in the body.
A number of methods exist for synthesizing L-2-oxothiazolidine-4-carboxylate and its carboxylic acid. Kaneko et al., Bull. Chem. Soc. (Japan) 37: 242-44 (1964), discloses that L-2-oxothiazolidine-4-carboxylate and its carboxylic acid can be synthesized by reacting L-cysteine with phosgene. Phosgene, however, is highly toxic and hazardous to use. Boettcher et al., Methods in Enzymology, 113(55): 458-60 (1985), discloses an improved method for synthesizing L-2-oxothiazolidine-4-carboxylate and its carboxylic acid, which uses phenyl chloroformate as a starting material instead of phosgene. The method disclosed in Boettcher et al. also results in higher product yields of L-2-oxothiazolidine-4-carboxylic acid than obtained when phosgene is used.
The reaction scheme of Boettcher et al. is generally as follows: ##STR1##
Generally, Boettcher et al. discloses that L-cysteine in an aqueous solution of potassium hydroxide and disodium ethylenediamine tetraacetate (Na.sub.2 EDTA) is reacted with additional potassium hydroxide and a mixture of phenyl chloroformate and toluene. This reaction yields the desired compound, sodium or potassium L-2-oxothiazolidine-4-carboxylate, as well as products consisting of the potassium and sodium salts of phenol and chloride. The solution is filtered to remove any particulate solids. The toluene layer is also removed. The aqueous layer or phase remaining, which contains L-2-oxothiazolidine-4-carboxylate and the salts of phenol and chloride, is acidified to a pH of about 6 to about 7 with hydrochloric acid so as to convert the potassium phenolate into phenol and additional potassium chloride.
The aqueous solution containing phenol, L-2-oxothiazolidine-4-carboxylate and potassium chloride is then contacted with diethyl ether in a staged manner to extract phenol from the aqueous solution into the immiscible diethyl ether phase. Boettcher et al. discloses that this extraction step is performed three times. The diethyl ether phase containing phenol is then separated from the aqueous phase containing L-2-oxothiazolidine-4-carboxylate. Thereafter the pH of the aqueous phase is further lowered to a pH of about 1 to about 2 by adding hydrochloric acid. This second pH adjustment converts the carboxylate to the carboxylic acid.
In Boettcher et al., L-2-oxothiazolidine-4-carboxylic acid is extracted from the aqueous solution with ethyl acetate which allows the potassium chloride to remain in the aqueous layer. The ethyl acetate layer is removed from the aqueous layer and the ethyl acetate is removed by evaporation. The resulting solid is dissolved in hot water and the L-2-oxothiazolidine-4-carboxylate is crystallized from this solution and dried. This product will be sieved for the correct particle size and packaged.
The addition of hydrochloric acid before and after extraction as disclosed in Boettcher et al. will add additional chloride ions to the aqueous layer containing L-2-oxothiazolidine-4-carboxylate to those already present in the solution as a normal byproduct of the chemical reaction between L-cysteine and phenyl chloroformate. Consequently, it is necessary to remove the original and additional potassium chloride to obtain the desired product.
The extraction and pH adjustment steps of the Boettcher et al. method are difficult to implement. The present invention greatly simplifies that method since the sequence of extraction steps utilizing diethyl ether or other extracting agent is replaced by contacting the reaction mixture in one step with an adsorption resin (however, multiple recycle steps can be used). The adsorption resin adsorbs at least about 90% and preferably about 95% to about 98% of the weight of phenol in the mixture and at most about 5%, preferably about 0% to about 3% of the weight of L-2-oxothiazolidine-4-carboxylate in the mixture. The improvement is also safer and less toxic than the Boettcher et al. method since it eliminates the need to use volatile and explosive extraction reagents such as diethyl ether. These benefits lead to a decrease in production cost when the process is carried out on a commercial scale.
The prior art discloses the removal of phenol by adsorption with an adsorption resin in applications other than the removal of phenol from a reaction mixture containing L-2-oxothiazolidine-4-carboxylate. For example, Mijangos et al., J. Chem. Eng. Data 40: 875-879 (1995) and Goto et al., Environ. Sci. Technol. 20: 463-467 (1986) discuss the adsorption of phenol using adsorption resins in the context of water pollutants. These applications, however, involve the removal of low concentrations of phenol. In the present invention, the concentration of phenol produced by the reaction is very high when compared to the concentration of the L-2-oxothiazolidine-4-carboxylate (about 0.64 kg phenol per 1.0 kg of compound). Based upon the teachings of these prior art references and upon the general concept that adsorption is used for removing materials of low concentration, one of ordinary skill in the art would not consider using adsorption to remove phenol from a solution of such high concentration as obtained in the synthesis of L-2 oxothiazolidine-4-carboxylate as described above, or in a similar manner.